Wang Maoxian, Guo Zhanyun, Wang Shilong
College of Life Sciences and Technology, Tongji University, Shanghai, PR China.
Gene Expr. 2012;15(5-6):235-41. doi: 10.3727/105221613x13571653093286.
Hydrogen sulfide (H2S), as an endogenous signaling molecule in mammals, shows a variety of biological effects. Cystathionine gamma-lyase (CSE)/H2S pathway has been implicated in scavenging reactive oxygen species (ROS) in the mammalian cells. Therefore, we first investigated the regulatory effects of exogenously applied hydrogen peroxide (H2O2) on CSE expression in the mammalian cells. African green monkey kidney fibroblastlike cells (COS-7 cells) or human embryonic kidney 293 cells (HEK 293 cells) were transfected with CSE promoter-luciferase reporter constructs and treated with H2O2 of 1, 5, and 10 microM for 0.5 and 1.5 h at 37 degrees C. The transfected cells were assayed for firefly luciferase activities normalized by Renilla luciferase activity. Human lung adenocarcinoma cells (A549 cells) or human liver cancer cells (SMMC-7721 cells) were treated with H2O2 of 1, 5, and 10 microM for 0.5 and 1.5 h at 37 degrees C, and were then harvested and analyzed by Western blotting and quantitative RT-PCR. Our results showed that the treatment of a medium concentration (5 microM) of H2O2 at a longer time (1.5 h) upregulated CSE expression in the mammalian cells at the levels of the promoter, message RNA, and protein. Collectively, exogenously applied H2O2 can not only markedly affect CSE mRNA and protein expression, but also can affect the CSE promoter activity in the mammalian cells. Our observations indicate that that exogenous H2O2 can upregulate the expression of the CSE gene in the mammalian cells, which will provide the possibility of the scavenging effect of the CSE gene indirectly on ROS in the mammalian cells. However, the regulatory mechanism involved in the effects of exogenously applied H2O2 on CSE expression in the mammalian cells need be further studied.
硫化氢(H₂S)作为哺乳动物体内的一种内源性信号分子,具有多种生物学效应。胱硫醚γ-裂解酶(CSE)/H₂S途径参与了哺乳动物细胞内活性氧(ROS)的清除。因此,我们首先研究了外源性过氧化氢(H₂O₂)对哺乳动物细胞中CSE表达的调控作用。将非洲绿猴肾成纤维样细胞(COS-7细胞)或人胚肾293细胞(HEK 293细胞)用CSE启动子-荧光素酶报告基因构建体转染,并在37℃下用1、5和10微摩尔的H₂O₂处理0.5小时和1.5小时。对转染后的细胞进行检测,以海肾荧光素酶活性标准化萤火虫荧光素酶活性。将人肺腺癌细胞(A549细胞)或人肝癌细胞(SMMC-7721细胞)在37℃下用1、5和10微摩尔的H₂O₂处理0.5小时和1.5小时,然后收获细胞并通过蛋白质免疫印迹法和定量逆转录-聚合酶链反应进行分析。我们的结果表明,在较长时间(1.5小时)用中等浓度(5微摩尔)的H₂O₂处理可上调哺乳动物细胞中启动子、信使RNA和蛋白质水平的CSE表达。总的来说,外源性应用的H₂O₂不仅能显著影响CSE的mRNA和蛋白质表达,还能影响哺乳动物细胞中CSE启动子的活性。我们的观察结果表明,外源性H₂O₂可上调哺乳动物细胞中CSE基因的表达,这将为CSE基因间接清除哺乳动物细胞内ROS的作用提供可能性。然而,外源性应用的H₂O₂对哺乳动物细胞中CSE表达影响的调控机制仍需进一步研究。